Genes in skin wound healing: microRNAs 2

Published on October 7, 2014   33 min
0:04
Now I'd like to switch gears and address another novel concept that has originated based on work in our laboratory. The basic premise here is that some elements of inducible wound healing is actually retained in the adult skin tissue in a silenced form. This silencing is primarily executed by microRNA. As one example, we have discovered that miR-200b in skin, microvascular endothelial cells, act as an angiostatic agent during wounding. This silencer is desilenced, enabling wound angiogenesis to start. Let's look at the data here. In Panel A, you see human microvascular dermal endothelial cells, a scratch assay has been done to study the migration of these cells. And if you added miR-200b mimic to these cells you clearly blunt or decelerate the migration of the cells across the scratch. In Panel B, it is shown that the ability of these cells to from tubes in a matrigel, which is a sign of angiogenic property, such angiogenic property is remarkably blunted or impaired in the presence of miR-200b mimic. Supporting the notion that miR-200b has potent angiostatic properties.
1:40
In the intact adult skin, it is critically important to keep angiogenesis under check. And miR-200b is one such silencer that contains angiogenic response under intact skin conditions. Following wounding, however, miR-200b levels in the skin sharply go down. In Panel A, we see that following 3 days after wounding miR-200b levels in the wound edge tissue is significantly lower. And as the wound heals, miR-200b comes up, and the expectation here is that this rebound of miR-200b expression is aimed at restoring homeostasis in the skin, meaning stopping unwanted additional angiogenesis after the wound has closed. Now this lowering of miR-200b following wounding, as noted in the wound test tissue in Panel A, what are the cell types primarily responsible for this type of lowering? The answer to that question, the wound test tissue was taken, and endothelial cells were laser captured from that wound test issue. And the results, as shown in Panel B, demonstrate that specifically endothelial cell, microRNA-200b, is substantially lowered after wounding.
3:10
To test the significance of miR-200b in wound angiogenesis, we sought to deliver the miR-200b gene to the skin. As you see in Panel A, the lentiviral gene delivery process resulted in almost doubling of the expression of 200b in the skin. Following such successful delivery of miR-200b to the skin, the skin was then wounded and angiogenic outcomes, such as blood flow and endothelial cell density, was enumerated. Panel B shows that 7 days after wounding the skin that was subjected to over expression of miR-200b, blood flow in the skin affected by the wound was substantially lower in response to miR-200b over expression. Similarly, it was noted that CD 31 positive endothelial cell count in the wound test tissue was also significantly lower. Suggesting that miR-200b does play an important role in resisting angiogenesis in the skin.
4:28
Diabetic wounds are subjected to a number of complications. Of which, one critical factor is excessive inflammation. We have demonstrated that high TNF levels during this condition of excessive inflammation associated with diabetic wounds, resist the lowering of miR-200b following wounding. As a result, even after wounding, miR-200b levels stay high. And that is one factor that impairs diabetic wound angiogenesis. We have also reported that sequestering this additional or excessive TNF alpha in the diabetic wound, helps bring down the miR-200b. As those of the which, wound angiogenesis may be jump started. Let us look at some of the data.
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Genes in skin wound healing: microRNAs 2

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